间充质干细胞对巨噬细胞活化及其功能影响的实验研究-内科血液病学专业论文.docxVIP

兰州大学硕士学位论文间充质干细胞对巨噬细胞活化及其功能 兰州大学硕士学位论文 间充质干细胞对巨噬细胞活化及其功能 影响的实验研究 中文摘要 目的探讨在体外条件下骨髓间充质干细胞(MSCs)对脂多糖(LPS)刺激后小 鼠腹腔巨噬细胞活化及其功能的影响。 方法采用贴壁筛选法分离、纯化BALB／e小鼠骨髓MSCs，用经过高压灭 菌的4％巯基乙酸钠溶液腹腔注射刺激，4天后收集小鼠腹腔巨噬细胞，依材料 和方法所述分组，建立MSCs与巨噬细胞共培养体系。用LPS(终浓度lug／m1) 刺激巨噬细胞18小时后收集细胞培养上清，检测培养体系中肿瘤坏死因子．a (TNF．Q)，转化生长因子．13(TGF．13和一氧化氮(NO)等炎症性细胞因子分 泌量的变化，同时在培养体系中加入大肠杆菌标准菌株(ATCC25922)，共孵 育24小时后用缓冲液洗涤，尽量除去未被吞噬的细菌后固定，采用瑞氏染色检 查巨噬细胞吞噬功能的变化。 结果 用巯基乙酸钠溶液腹腔注射刺激方法收集的小鼠腹腔巨噬细胞为炎 症性巨噬细胞，其培养上清中可检测到少量细胞因子，经LPS进一步刺激活化 后培养上清中TNF．Q和NO的含量明显上升，分别为(147．414-37．13)pg／ml， (59．934-8．74)uM；而与MSCs共培养时，TNF—a显著减少【(97．58+30．26)pg／ml， P=0．0321，NO降N(50．904-29．48)uM，P0．051当培养体系中存在MSCs上清时， 即MPMs+LPS+MSC上清组中，TNF—a和NO进一步减少为(58．28+3 1．54)pg／ml， 【．3．36-A：2．31)uM，P0．0005；然而，各组中TGF-13含量的差异无统计学意义，P O．05。MSCs对巨噬细胞活化后的吞噬率及吞噬指数没有影响。 结论实验首次发现，MSCs可抑制LPS刺激后小鼠腹腔巨噬细胞的活化， 而对其吞噬功能没有影响。 关键词骨髓间充质干细胞；巨噬细胞；活化；脂多糖 兰州大学硕士学位论文Effect 兰州大学硕士学位论文 Effect of bone marrow mesenchymal stem cells 0n activation and lunction oI mouse oeritoneal macropnages Abstract objeetive To explore the influence of bone marrow mesenchymal stem cells (MSCs)on maerophage activation after lipopolysarchride stimulation． Methods BALB／c mouse bone marrow MSCs were generated using general method，mice were injected intrapefitoneally、Ⅳim 1 ml of 4％sterile Brewer TG medium 4 days prior to harvest of the macrophages by peritoneal lavage．TG-elicited macrophages were prepared as the plastic tissue culture plate-adherent population of cells from peritoneal exudates lavage flued essentially,nonadherent cells were removed by gentle washing,and the remaining adherent cells were cultured for experiment．The CO-culture system Was established by planting macrophages on the MSCs monolayer．The supernatants were collected for TNF·a／TGF·B and NO assay after Stimulming macrophages、析tll LPS for about 1 8 hours．At the same time Escbericbia coli standard strain(ATCC25922)was added into the culture system and incubated for another 24 hours，macrophages were stained and the changes of phagocytosis were e